A knowledge of the regulation of gene activity is central to our understanding of processes of growth and differentiation in normal and malignant cells. Most mammalian cells are now known to contain multiple forms of the enzyme, DNA-dependent RNA polymerase. These isozymes are intimately involved in regulating the type and amount of RNA synthesized in the nucleus of the cell. These enzyme activities may therefore play a central role in the phenotypic expression of cells. The proposed study will extend our efforts to investigate the transcriptional activity, structure and metabolism of the multiple forms of this enzyme in normal and malignant mouse tissues. Present efforts are directed to examining the RNA polymerase profiles in normal mouse liver and in a variety of mouse tumors to determine whether the pattern changes qualitatively as well as quantitatively in tumor cells. At the same time, efforts will be made to isolate sufficient of the major forms of the enzyme from these tissues for meaningful studies of their structure, intranuclear localization and metabolism. Many of the analytical and preparative procedures are centered around our method for separating these isozymes by gel electrofocusing. This system allows the simultaneous fractionation of small amounts of tissue and displays in one step and at high resolution the various isozymic forms in crude nuclear extracts. It also has unique advantages for preparative purposes. The subunit composition and structural relationships of the major forms will be investigated by biochemical and immunological procedures. Rates of synthesis and turnover of the isozymes and their constituent subunits will be investigated by tracer studies in vivo and in cell grown in tissue culture. Attempts will be made to elucidate molecular mechanisms leading to alterations in the activities of the polymerase isozymes at different stages of growth and development. In parallel studies, the intranuclear disposition of the isozymes at different stages of cell growth will be investigated with fluorescent antibody techniques.